Breast tumors may obtain the fatty acids they require for growth and survival by two distinct mechanisms. Lipids may be produced de novo (lipogenesis) by tumor cells that express lipogenic enzymes, a process supported by nuclear protein S14. Alternatively, fatty acids may be cleaved from lipoproteins in the blood (lipolysis) by the extracellular enzyme lipoprotein lipase (LPL). Breast cancer cells do not make LPL, but adipocytes of the mammary fat pad supply it locally. During metastasis, however, tumor cells encounter environments such as lymph node and the general circulation that are devoid of LPL, and must depend on lipogenesis. We hypothesize that S14 1) promotes enhanced lipid synthesizing capacity that is crucial during breast cancer metastasis, and 2) that provision of LPL by adjacent tissue promotes growth of primary and metastatic breast cancers, particularly those with low S14 and lipogenic capacity. The hypothesis rests on observations that S14 is overexpressed in most breast cancers, S14 drives growth and survival of cultured breast cancer cells, S14 overexpression predicts breast cancer recurrence, and breast cancer cells do not express LPL. Our Aims focus on interactions of S14 and local availability of LPL in a murine breast cancer model, and on mechanisms of S14 action. Mice with mammary epithelial-specific S14 gene knockout will be bred with animals lacking LPL in the primary tumor site (mammary gland) and the major metastatic site (lung), and the MMTV-Her2 model. Tumor incidence, growth, metabolism, metastasis, and angiogenesis will be assessed. First we will define the roles of S14 and local LPL by comparing the S14 mammary knockout and tissue-restricted LPL mutations alone and together in the Her2 model. As dietary fat substrate will condition the impact of altered LPL, effects of low- or high-fat diets will be assessed. Second, we will assess the impact of enforced mammary S14 overexpression. Third, we will identify proteins that interact with S14 in breast cancer cells, and define structural requirements for assembly of S14 multimers. This will provide proof of principle for targeting S14 in breast cancer, identify vulnerable points of the molecule to achieve it, and provide a novel, comprehensive model of tumor lipid metabolism that takes lipogenesis, lipolysis, and diet into account. Lay Summary: S14 is found in aggressive breast cancers, where it promotes formation of fats they require for growth. We will study breast cancer-prone mice with high or absent mammary expression of S14 fed high and low fat diets, and will also use a biochemical approach to determine how S14 works. This work will pave the way for anti-S14 treatments for breast cancer patients.